Apparatus for washing particle matter



NOV. 14, 1967 51555 3,352,486

APPARATUS FOR WASHING PARTICLE MATTER Filed A ril 30, 1965 2 etsh 1VACUUM SOURCE .9

mygmoa Mary B. Gibbs ATTORNEY Nov. 14, 1967 M. B. GIBBS 3,352,436

APPARATUS FOR WASHING PARTICLE MATTER.

Filed April 30, 1965 -2 Sheets-Sheet 2 5 FIG.3

' FIG 5' 7/, t T 2o 18 22 i J 23 I 23 1 FlG.-2

T T 14 l l 14 mvrsmoa 'Mary B. Gibbs BY "@1 79, MM-

ATTORNEY United States Patent 3,352,486 APPARATUS FOR WASHING PARTICLEMATTER Mary B. Gibbs, Hyattsville, Md., assignor to Clay-Adams, Inc.,New York, N.Y., a corporation of New York Filed Apr. 30, 1965, Ser. No.452,320 6 Claims. (Cl. 233-26) This invention relates to methods andapparatus for washing particle matter; and more particularly concernsprocedures and means for washing erythrocytes and other formed elementsof the blood free of contaminating serum proteins or other material.

conventionally, erythrocytes are washed by first centn'fuging the bloodsample in a test tube to pack the red cells, decanting the supernatantplasma or serum, adding washing solution, inverting and shaking the testtube manually, centrifuging and decanting. This may be repeated, withsubsequent additions of wash solution, several times, to achieve thedesired degree of removal of contaminating agents. However,contamination may occur due to the tendency of droplets to cling to thetest tube wall, leaving residual matter on the test tube wall. Also,decantation may deliberately be incompletely carried out in order toavoid loss of cells.

An object of this invention is to provide improved methods and apparatusfor washing centrifuged blood samples and eliminating the temptation toseal test tubes with finger tips while resuspending the sediment. Thelatter practice is known to cause cross contamination from specimen tospecimen.

Another object of this invention is to reduce the danger of loss ofsediment when supernatant material is hastily decanted.

A further object of this invention is to provide improved apparatus forsimultaneously washing a large number of blood samples in individualtest tubes while such tubes are still mounted in a centrifuge, after thecentrifuging operation, thereby avoiding inadvertent mistakes inidentification of the individual specimens, not uncommon in known manualwashing procedures.

Another object of this invention is to provide improved transfer meansfor supplying washing solutions to blood samples carried in test tubesbefore centrifugation, the transfer means also serving to removesupernatant material after centrifugation of the samples; the transfermeans further serving to supply Washing solution in a manner to dislodgethe cells from the tube walls and to carry the formed elements into theupper portions of the tubes.

A further object of this invention is to provide manifold type, improvedmeans for injecting washing solution into test tubes carrying bloodsamples and for removing super natant material without decantation so asto materially reduce danger of cellular loss when the washing operationis repeated; I

Still another object of this invention is to provide improved washingapparatus in manifold form for ejecting washing solution into individualtest tubes carrying centrifuged blood specimens, the ejected stream ofwashing solution being located and directed in a manner to dislodge thewashed, centrifuged pack of red cells from the tube wall in an efiicientmanner.

Known, manual procedures for washing cells does not permitstandardization of the cell washing operation. Accordingly, yet anotherobject of this invention is to provide a cell washing procedure andapparatus which permits standardization of the same, thereby producing awashed cell residue of a given dryness.

Other objects of this invention will in part be obvious and in parthereinafter pointed out.

In the drawings:

FIG. 1 is a side elevational view of apparatus for washing blood samplesand embodying the invention herein;

FIG. 2 is a perspective view of the sample carrying tubes disposed inthe centrifuge basket;

FIG. 3 is a perspective view showing the manifold Vdevice for ejectingand aspirating washing solution, embodying the invention;

" stant invention comprises a manifold head arranged for connection to asource of washing solution which may be delivered to the head under agiven'pressure for distribution to individual dispensing tubesconveniently arranged in a manner, whereby the dispensing tubes may beinserted into the test tubes carrying the blood specimens to be washedas said test tubes are disposed in a suitable arrangement on the head ofa conventional centrifuge. The manifold head may also be subjected tonegative pressures whereby supernatant material from the test tubes maybe aspirated without unduly disturbing the centrifuged deposit of redcells. Also, the dispensing tubes extending from the manifold head areso located in respect to the test tube walls as to effectively dislodgethe centrifuged deposit of cells, when further washing solution ispassed to the dispensing tubes under pressure.

Thus, as shown in FIG. 1, 10 designates a conventional centrifuge of thetype used in serology and which carries within the upper portion thereofthe usual test tube basket 11, see FIG. 2, which includes slotted discs12, 13 for locating the individual test tubes T, which carry the bloodspecimens to be washed, in circular arrangement. The discs 12, 13 areformed with radial slots 14, 15 respectively to allow the tubes T toassume inclined positions during the centrifuging operation and to bedisposed vertically before and after such centrifuging operation.

Means is provided for delivering washing solution, such asv the usualsaline solution, simultaneously to each test tube T while such tubes Tare still carried in basket 11 in centrifuge 10. To this end, there isprovided a manifold head 16 in the form of a flat circular hollow memberhaving a top wall 17 and having a centrally located opening 18 thereinfrom which extends an upstanding connector, tubular member 19, and abottom wall 20, thus forming a circular chamber 21 which is incommunication with tube 19. The bottom wall 20 of chamber 21 is formedwith a circular arrangement of openings 22 disposed adjacent theperiphery thereof. Small bore tubings 23 are fitted at their upper endsinto openings 22 and are thus in communicating relation with chamber 21.

The head 16 is adapted to be disposed in aligned relation to test tubesT so as to locate each tubing 23 over a test tube T, for insertion andwithdrawal relative thereto. When tubings 23 are fully inserted intotest tubes T, the tips thereof are located adjacent the lower ends ofthe test tubes.

A collar 24 formed with through openings 24A arranged in a circle isadapted to be slidably mounted on tubes 23, chain means 25interconnecting head 16 with collar 24 to limit the lowered movement ofsaid collar on tubings 23.

As shown in FIG. 1, upstanding tube 19 on head 16 is adapted to beconnected to a supply of washing solution as may be carried in acontainer 26, by way of a flexible tubing 27, means for deliveringmeasured quantities of washing solution, such as saline, as by ahypodermic syringe 28 and vflexible tubing 29 connected to connectormember 19.

In use, the manifold head 16 is located over the test tubes T while theyare in their vertical positions in centrifuge the movable collar 24facilitating the proper vertical positioning of tubings 23 for entryinto the respective test tubes T.

Syringe 28 is operated to withdraw a selected amount of washing solutionfrom container 26 to chamber 21 of the head 16 and thence to tubings 23.

The manifold 16 is then removed from its dispensing position to allowthe cover of centrifuge 10 to be closed and the centrifuge to beoperated in the usual manner. After centrifugation has been completed,supernatants in each of test tubes T may be removed without decantation,by means of a second manifold head 16A entirely similar to thatpreviously described, except that its flexible tubing 29A is connectedto a source of reduced pressure as a vacuum pump 30 and a receiver 31.

The blood samples in test tubes T may be repeatedly washed with salinewashing solution from container 26 by repeating the operations describedabove; the syringe 28 being retracted to fill the same with new chargesof washing solution in measured amount which is ejected into dispensingtubes 23 when the plunger of syringe 28 is pressed forwardly.

When the supernatants are aspirated out of the test tubes T after theprevious washing operation, fresh washing solution is ejected fromtubings 23 by appropriate movement of the syringe plunger and thedeposited red cells C or other formed elements collected on the outerwall portion 32 or bottom wall portion 33 of each of the test tubes Tdue to successive centrifugation, will be dislodged and carried to theupper portions of tubes T in the wash solution. The washing cycle may berepeated two or more times with intermediate short periodcentrifugation. The washed cells may then be used in conventional testprocedures.

The forced concentration of formed elements in the upper portion of testtubes T is an extremely effective washing operation inasmuch as thegreater distance be tween the centroid of the formed elements and thebottom of said test tubes allows for more contact between the washingsolution and each particle during centrifugation.

It is understood that the Washing operation may be carried out with asingle manifold head 16, using the same for dispensing washing solutionand for aspirating supernatants in alternation. However, in such case,the head 16, its dispensing tubes 23 and connecting flexible tubing 29are well rinsed between such operations to avoid any possibility ofcontamination.

While the manifold head 16 and its tubes 23 are shown as particularlyadapted for use with specimen tubes normally in a vertical position, itis understood that such head may be modified to dispose itsdispensing-aspirating tubes at a selected angle to correspond to thedisposition of test tubes arranged at a corresponding angle.

While the apparatus and process herein disclosed is shown in connectionwith the washing of erythrocytes, it is understood that the same may beused in washing other particle matter such as latex, bentonite,bacteria, spores, tissue cells, chemical and bacteriologicalprecipitates.

What is claimed is:

1. Cell washing means for use with a plurality of test tubes, each testtube carrying a centrifuged blood specimen portion deposited at thelower end thereof, said 4 washing means comprising means for supportingsaid test tubes in an upstanding arrangement, manifold means movabletowards and away from said supporting means, said manifold meanscomprising a plurality of transfer tubings depending therefrom and incommunication therewith, said transfer tubings being in a verticalarrangement, said manifold means being movable toward said supportingmeans with said transfer tubings in respective alignment with saidsupported test tubes for introducing said transfer tubings into therespective test tubes, said tubings having a length to locate the lowerends thereof at a point within said test tubes adjacent the deposittherein, said manifold means including inlet means for admitting washingsolution therein and the distribution thereof via said transfer tubingsto the lower end portion of each test tube for washing the deposittherein.

2. Cell washing means as in claim 1 wherein said inlet means on themanifold means is adapted to be con.- nected to a source of reducedpressure whereby to aspirate supernatants from each of said test tubesby way of said transfer tubings and passages in said manifold means.

3. Cell washing means as in claim 1 wherein each of said transfertubings comprises small bore tubing, and collar means having a pluralityof through openings arranged for passing the respective tubingstherethrough, said collar means being slidably mounted on said tubingsfor movement to positions to properly locate the lower portions of saidtubings for entry into the respective test tubes.

4. For use with test tubes carrying blood specimens to be subjected towashing solutions and centrifugation to effect the separation of liquidfrom the formed elements thereof, said test tubes being in a centrifugehead, improved cell washing means comprising a pair of similar manifoldheads, movable towards and away from said centrifuge heads, eachmanifold head comprising a chamber and inlet means communicating withsaid chamber, and a plurality of transfer tubings mounted on saidmanifold head, said tubings communicating at one end thereof with saidchamber, the arrangement of tubings on said manifold heads beingcorrelated to the arrangement of the test tubes on said centrifuge toallow the tubings to be respectively inserted into the test tubes witheach tubing lying within the test tube associated therewith, saidtubings having a length to locate the outer ends thereof adjacent thelower ends of said test tubes when said tubings are fully inserted intosaid test tubes, one of said heads having the inlet means thereofarranged for connection to a source of washing solution, the other ofsaid heads having the inlet means thereof arranged for connection to asource of reduced pressure whereby said heads may be used in successionto (1) inject washing solution into each of said test tubes; and (2) toaspirate supernatants from each of said test tubes.

'5. Cell washing means as in claim 4 wherein the tubings on saidmanifold head are in a circular arrangement.

6. In combination with a manifold comprising a chamber portion inletmeans on said manifold in communication with said chamber portion and aplurality of flexible transfer tubings mounted on said manifold and incommunication with the chamber portion thereof, said tubings beingarranged in a predetermined pattern and in parallel relation to eachother, means for maintaining the outer ends of said tubings in parallelrelation to each other comprising collar means having a plurality ofthrough openings of a diameter for slidably passing the respectivetubings therethrough, whereby said collar means may be slidably movedrelative to said tubings toward the outer ends thereof to bring theouter ends of said tubings into parallel relation to each other.

(Other references on following page) 6 References Cited 3,202,348 8/1065Strohmaier 23326 UNITED STATES PATENTS 3,208,468 9/1965 Hansen 137-2661/1937 Lux 23272.7 1 014 348 8 igfi Gq PATENTS 8/1956 Hatcher 239 229 5ermany' 10/ 1958 Snyder 23-272] HENRY T. KLINKSIEK, Primary Examiner.

2/1965 Berg 239--229 X M. CARY NELSON, Examiner.

1. CELL WASHING MEANS FOR USE WITH A PLURALITY OF TEST TUBES, EACH TESTTUBE CARRYING A CENTRIFUGED BLOOD SPECIMEN PORTION DEPOSITED AT THELOWER END THEREOF, SAID WASHING MEANS COMPRISING MEANS FOR SUPPORTINGSAID TEST TUBES IN AN UPSTANDING ARRANGEMENT, MANIFOLD MEANS MOVABLETOWARDS AND AWAY FROM SAID SUPPORTING MEANS, SAID MANIFOLD MEANSCOMPRISING A PLURALITY OF TRANSFER TUBINGS DEPENDING THEREFROM AND INCOMMUNICATION THEREWITH SAID TRANSFER TUBINGS BEING IN A VERTICALARRANGEMENT, SAID MANIFORLD MEANS BEING MOVABLE TOWARD SAID SUPPORTINGMEANS WITH SAID TRANSFER TUBINGS IN RESPECTIVE ALIGNMENT WITH SAIDSUPPORTED TEST TUBES FOR INTRODUCING SAID TRANSFER TUBINGS INTO THERESPECTIVE TEST TUBES, SAID TUBINGS HAVING A LENGTH TO LOCATE THE LOWERENDS THEREOF AT A POINT WITHIN SAID TEST TUBES ADJACENT THE DEPOSIT